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- Hellström, Mats, 1976, et al.
(författare)
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Bioengineered uterine tissue supports pregnancy in a rat model
- 2016
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Ingår i: Fertility and Sterility. - : Elsevier BV. - 0015-0282. ; 106:2
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Tidskriftsartikel (refereegranskat)abstract
- Objective: To create a bioengineered uterine patch for uterine repair of a partially defect uterus. Design: Three different decellularized uterine scaffolds were recellularized in vitro with primary uterine cells and green fluorescent protein-(GPF-) labeled bone marrow-derived mesenchymal stem cells (GFP-MSCs). The patches were transplanted in vivo to investigate their tissue adaptation and supporting capacity during pregnancy. Animal(s): Female Lewis rats (n = 9) as donors to generate whole-uterus scaffolds using three different protocols (n = 3 per protocol); Sprague Dawley rats (n = 40) for primary uterus cell isolation procedures (n = 10) and for transplantation/pregnancy studies (n = 30); and male Sprague Dawley rats (n = 12) for mating. Intervention(s): Decellularization was achieved by whole-uterus perfusion with buffered or nonbuffered Triton-X100 and dimethyl sulfoxide (DMSO; group P1/P2) or with sodium deoxycholate (group P3). Primary uterine cells and GFP-MSCs were used to develop uterine tissue constructs, which were grafted to uteri with partial tissue defects. Main Outcome Measure(s): Recellularization efficiency and graft quality were analyzed morphologically, immunohistochemically, and by real-time quantitative polymerase chain reaction (PCR). The location and number of fetuses were documented during pregnancy days 16-20. Result(s): Pregnancy and fetal development were normal in groups P1 and P2, with fetal development over patched areas. Group P3 showed significant reduction of fetal numbers, and embryos were not seen in the grafted area. Quantitative PCR and immunohistochemistry revealed uterus-like tissue in the patches, which had been further reconstructed by infiltrating host cells after transplantation. Conclusion(s): Primary uterine cells and MSCs can be used to reconstruct decellularized uterine tissue. The bioengineered patches made from triton-X100+DMSO-generated scaffolds were supportive during pregnancy. These protocols should be explored further to develop suitable grafting material to repair partially defect uteri and possibly to create a complete bioengineered uterus. ((C) 2016 by American Society for Reproductive Medicine.)
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| 2. |
- Oltean, Mihai, 1976, et al.
(författare)
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Optimal Solution Volume for Luminal Preservation: A Preclinical Study in Porcine Intestinal Preservation
- 2016
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Ingår i: Transplantation Proceedings. - : Elsevier BV. - 0041-1345. ; 48:2, s. 532-535
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Tidskriftsartikel (refereegranskat)abstract
- Background. Rodent studies suggest that luminal solutions alleviate the mucosal injury and prolong intestinal preservation but concerns exist that excessive volumes of luminal fluid may promote tissue edema. Differences in size, structure, and metabolism between rats and humans require studies in large animals before clinical use. Methods. Intestinal procurement was performed in 7 pigs. After perfusion with histidine-tryptophan-ketoglutarate (HTK), 40-cm-long segments were cut and filled with 13.5% polyethylene glycol (PEG) 3350 solution as follows: VO (controls, none), V1 (0.5 mL/cm), V2 (1 mL/cm), V3 (1.5 mL/cm), and V4 (2 mL/cm). Tissue and luminal solutions were sampled after 8, 14, and 24 hours of cold storage (CS). Preservation injury (Chiu score), the apical membrane (Z0-1, brush-border maltase activity), and the electrolyte content in the luminal solution were studied. Results. In control intestines, 8-hour CS in HTK solution resulted in minimal mucosal changes (grade 1) that progressed to significant subepithelial edema (grade 3) by 24 hours. During this time, a gradual loss in ZO-1 was recorded, whereas maltase activity remained unaltered. Moreover, variable degrees of submucosal edema were observed. Luminal introduction of high volumes (2 mUmL) of PEG solution accelerated the development of the subepithelial edema and submucosal edema, leading to worse histology. However, ZO-1 was preserved better over time than in control intestines (no luminal solution). Maltase activity was reduced in intestines receiving luminal preservation. Luminal sodium content decreased in time and did not differ between groups. Conclusions. This PEG solution protects the apical membrane and the tight-junction proteins but may favor water absorption and tissue (submucosal) edema, and luminal volumes >2 mL/cm may result in worse intestinal morphology.
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| 3. |
- LeVaillant, C. J., et al.
(författare)
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Significant changes in endogenous retinal gene expression assessed 1 year after a single intraocular injection of AAV-CNTF or AAV-BDNF
- 2016
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Ingår i: Molecular Therapy - Methods and Clinical Development. - : Elsevier BV. - 2329-0501. ; 3
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Tidskriftsartikel (refereegranskat)abstract
- Use of viral vectors to deliver therapeutic genes to the central nervous system holds promise for the treatment of neurodegenerative diseases and neurotrauma. Adeno-associated viral (AAV) vectors encoding brain-derived neurotrophic factor (BDNF) or ciliary derived neurotrophic factor (CNTF) promote the viability and regeneration of injured adult rat retinal ganglion cells. However, these growth-inducing transgenes are driven by a constitutively active promoter, thus we examined whether long-term AAV-mediated secretion of BDNF or CNTF affected endogenous retinal gene expression. One year after the intravitreal injection of AAV-green fluorescent protein (GFP), bi-cistronic AAV-BDNF-GFP or AAV-CNTF-GFP, mRNA was extracted and analyzed using custom 96 well polymerase chain reaction arrays. Of 93 test genes, 56% showed significantly altered expression in AAV-BDNF-GFP and/or AAV-CNTF-GFP retinas compared with AAV-GFP controls. Of these genes, 73% showed differential expression in AAV-BDNF versus AAV-CNTF injected eyes. To focus on retinal ganglion cell changes, quantitative polymerase chain reaction was undertaken on mRNA (16 genes) obtained from fixed retinal sections in which the ganglion cell layer was enriched. The sign and extent of fold changes in ganglion cell layer gene expression differed markedly from whole retinal samples. Sustained and global alteration in endogenous mRNA expression after gene therapy should be factored into any interpretation of experimental/clinical outcomes, particularly when introducing factors into the central nervous system that require secretion to evoke functionality. © 2016 Official journal of the American Society of Gene & Cell Therapy
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| 4. |
- You, S. W., et al.
(författare)
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Large-scale reconstitution of a retina-to-brain pathway in adult rats using gene therapy and bridging grafts: An anatomical and behavioral analysis
- 2016
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Ingår i: Experimental Neurology. - : Elsevier BV. - 0014-4886. ; 279, s. 197-211
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Tidskriftsartikel (refereegranskat)abstract
- Peripheral nerve (PN) grafts can be used to bridge tissue defects in the CNS. Using a PN-to-optic nerve (ON) graft model, we combined gene therapy with pharmacotherapy to promote the long-distance regeneration of injured adult retinal ganglion cells (RGCs). Autologous sciatic nerve was sutured onto the transected ON and the distal end immediately inserted into contralateral superior colliculus (SC). Control rats received intraocular injections of saline or adeno-associated virus (AAV) encoding GFP. In experimental groups, three bi-cistronic AAV vectors encoding ciliary neurotrophic factor (CNTF) were injected into different regions of the grafted eye. Each vector encoded a different fluorescent reporter to assess retinotopic order in the regenerate projection. To encourage sprouting/synaptogenesis, after 6 weeks some AAV-CNTF injected rats received an intravitreal injection of recombinant brain-derived neurotrophic factor (rBDNF) or AAV-BDNF. Four months after surgery, cholera toxin B was used to visualize regenerate RGC axons. RGC viability and axonal regrowth into SC were significantly greater in AAV-CNTF groups. In some cases, near the insertion site, regenerate axonal density resembled retinal terminal densities seen in normal SC. Complex arbors were seen in superficial but not deep SC layers and many terminals were immunopositive for presynaptic proteins vGlut2 and SV2. There was improvement in visual function via the grafted eye with significantly greater pupillary constriction in both AAV-CNTF + BDNF groups. In both control and AAV-CNTF + rBDNF groups the extent of light avoidance correlated with the maximal distance of axonal penetration into superficial SC. Despite the robust regrowth of RGC axons back into the SC, axons originating from different parts of the retina were intermixed at the PN graft/host SC interface, indicating that there remained a lack of order in this extensive regenerate projection. (C) 2016 Elsevier Inc. All rights reserved.
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